Visualizing cassava bacterial blight at the molecular level using CRISPR-mediated homology-directed repair

SUMMARY Research on a few model, plant-pathogen systems has benefitted from years of tool and resource development. This is not the case for the vast majority of economically and nutritionally important plants, creating a crop improvement bottleneck. Cassava bacterial blight (CBB), caused by Xanthomonas axonopodis pv. manihotis ( Xam ), is an important disease in all regions where cassava ( Manihot esculenta Crantz) is grown. Here we describe development of a tool for molecular-level visualization of CBB dynamics in vivo . Using CRISPR-mediated homology-directed repair (HDR), we generated plants containing scarless insertion of a GFP reporter at the CBB susceptibility ( S ) gene MeSWEET10a . Activation of MeSWEET10a-GFP by Xam was subsequently visualized at transcriptional and translational levels. Development of this tool was facilitated by a time-saving, adaptable strategy for identifying successful products of HDR, currently a limiting factor in plant research. This strategy has the potential to enable such research in other systems, improving the practicality of HDR-based experimentation.